Project Summary/Abstract : Natural products still comprise the majority of therapeutic agents. However, they suffer from a difficulty in the creation of derivatives, and they can also be hard to supply. Derivatives are extremely important in the optimization of properties, such as toxicity or solubility, which are critical to development of therapeutics. Recently, it has become increasingly popular to create libraries of derivatives using genetic engineering of natural product biosynthetic genes. Even in the best libraries, the numbers of derivatives are in the hundreds or low thousands. Recently, we discovered an evolutionary pathway that could potentially be harvested to generate up to billions of derivatives. The compounds can be rapidly "evolved" to improve desired properties while minimizing unwanted effects. We have developed a system in which new activities can be pharmacologically optimized in a matter of days to weeks, exploring a huge diversity of unnatural natural products. In this project, we will develop this system and provide a practical proof-of-concept using the potential anticancer natural product, Trunkamide, as a model. As a practical proof-of-concept, we will optimize the anticancer cyanobactin natural product, Trunkamide, which is a potential clinical agent that requires optimization. Our aims here are to: 1) Demonstrate the application of engineered cyanobactin libraries to high throughput screening and to problems of ADME/Tox. 2) Optimize the activity and pharmacological properties of a known pre-clinical anticancer candidate cyanobactin, Trunkamide, for clinical development. PUBLIC HEALTH RELEVANCE: Project Narrative: We will use innovative new technology to develop optimized anticancer agents with new mechanisms. The underlying technology to be developed has broad application in the treatment of human diseases, especially those involving infective agents or protein interactions.